Cell Biology/Signaling Diet-Induced Aortic Valve Disease in Mice Haploinsufficient for the Notch Pathway Effector RBPJK/CSL

Objective—Calcific aortic valve disease is similar to atherosclerosis in that both diseases result from chronic inflammation and endothelial dysfunction. Heterozygous NOTCH1 mutations have been associated to calcific aortic disease and a bicuspid aortic valve. We investigated whether mice with genetic inactivation of the Notch signaling pathway are prone to develop valve disease when exposed to a predisposing diet. Methods and Results—Using Doppler echocardiography, histology, immunohistochemistry, quantitative gene expression analysis, and cell culture assays, we examined the effect of a hypercholesterolemic diet supplemented with vitamin D on mice heterozygous for null mutations in the Notch1 receptor or the effector transcription factor gene RBPJk. After 16 weeks on the hyperlipidemic diet, calcific aortic disease was detected in heterozygous RBPJk mice. Analysis of valve leaflets revealed macrophage infiltration, enhanced collagen deposition, proosteogenic protein expression, and calcification. Heterozygous null Notch1 mice displayed milder histopathologic changes and did not develop any significant hemodynamic disturbance. Valvular disease correlated with reduced expression of the Notch target gene Hey1 in valves of RBPJk heterozygous mice fed the hyperlipidemic diet. Consistent with the in vivo data, Notch signaling inhibition in porcine valve interstitial cells led to downregulation of HEY1 transcription, activation of osteogenic markers, and increased calcified nodule formation. Conclusion—We show that Notch signaling disruption via RBPJk heterozygous inactivation results in aortic valve disease. Notch1 heterozygous mice do not show functional impairment, suggesting that additional Notch receptors may be involved in aortic valve homeostasis and disease. Our data establish a genetic mouse model of calcific aortic valve disease and may help to identify a patient population with reduced valvular NOTCH signaling at risk for developing this disease. C alcific aortic valve disease (CAVD) is a growing health problem in Western countries. Currently, more than 2% of people over the age of 65 and 4% over the age of 85 experience this condition, and the prevalence of this pathology is rising because the population is aging. 1 Recent research has shown that CAVD results from a complex inflammatory process similar to that found in atherosclerosis. In fact both diseases share common risk factors, such as age, smoking, hypertension, and hypercholesterolemia, involving common molecular mechanisms. 2 The Notch pathway is a local cellular signaling system that regulates cell fate determination, differentiation, and tissue patterning, and dysregulated Notch activity has pathological consequences. 3 Notch genes encode a group of transmembrane receptors (Notch1 to Notch4 in mammals), with a large extra-cellular region and …